Food Biochemistry and Food Processing (2 edition)

BLBS102-c35 BLBS102-Simpson March 21, 2012 14:9 Trim: 276mm X 219mm Printer Name: Yet to Come

35 Biochemistry and Probiotics 679

fats made this ingredients as a very good source of prebiotic

components for human body as well as yogurt and probiotics

bacteria (Boye et al. 2010). Enriching milk with prebiotic sup-

plements (Capela et al. 2006) enhances the stability of probiotics

in yogurt, but very few studies have been conducted with soy or

pulses. Addition of soy protein isolates in yogurt does not im-

prove stability during storage (Pham and Shah 2009) but some

pulses do (Zare et al. 2011).

Food fermentations with probiotics have mainly been con-

ducted on dairy and soy substrates. Pulses contain many of the

carbohydrates of soy but there are also various other oligosaccha-

rides (Table 35.3). Therefore, from a carbohydrate perspective,

growth of probiotics in pulses should require similar enzymatic

profiles to those required in soy; however, a wider range of cul-

tures can theoretically develop in some pulses due to the presence

of verbascose or other oligosaccharides.

Cereal-based products also offer many possibilities for the de-

velopment of probiotic-based (Farnworth 2004). Cereals mainly

have starch. Assimilation of starch typically requires amylases

and maltases.

Vegetables are also frequently used as substrates for lactic

acid fermentations and are thus potential matrices for probi-

otic cultures (Farnworth 2004). Sucrose is often found in these

matrices, but a wide variety of substrates are encountered. In cel-

ery, for example, mannitol is the main carbon-based substrate

but original polysaccharides are also discovered (Thimm et al.

2002).

It would take too long to examine each plant-based matrix

for the required enzymes. Following are the points that must be

emphasized:

1. Carbohydrate substrates vary as a function of the food
   matrix (Table 35.3) and, thus enzymatic requirements vary
   accordingly.


2. There is variability in the ability of probiotic bacteria to
   use the carbohydrates (Tables 35.1 and 35.2); a strain
   selection process is required.

BIOCHEMISTRY AND STABILITY

DURING STORAGE IN FOODS

Protection Against Oxygen

Probiotic bacteria are sensitive to many processes (heating,

freezing, aeration) associated with food production or storage

conditions (Champagne et al. 2005). But they are also sensi-

tive to some conditions that occur during storage. The two most

important are the presence of oxygen and high acidity of the

food. In both cases, some enzymatic systems may enhance the

survival of the cultures to these stressful conditions.

Since the intestines offer an anaerobic environment, many

strains are not adapted to growth in aerobic conditions. Techno-

logical adaptations can be made: microencapsulation, addition

of antioxidants, packaging conditions (Talwalkar and Kailasap-

athy 2004, Jimenez et al. 2008). But some strains do possess en-

zymatic systems that enhance their stability in foods containing

oxygen. The major problem associated with the presence of oxy-

gen is that hydrogen peroxide is produced in various metabolic

pathways. The two major enzyme systems that eliminate this

toxic by-product of oxygen are NADH oxidase and NADH per-

oxidase (Shimamura et al. 1992, Talwalkar and Kailasapathy

2003). These systems are inducible. Thus, when cultures are

gradually exposed to peroxide, an increased synthesis of these

enzymes occurs. NADH oxidase and NADH peroxidase function

optimally at pH 5.0 (Talwalkar and Kailasapathy 2003), which

suggests that the protection to oxygen would be higher in acid

foods (yogurt, fruit juices) than in neutral foods (unfermented

milk, vegetables). Not surprisingly, it was shown that when a

probiotic culture was added at the beginning of a yogurt fermen-

tation process, it was more stable during storage than when it

was added directly during the finished product (Hull et al. 1984).

One hypothesis was that the cultures were gradually exposed to

H 2 O 2 produced byL. delbrueckiissp.bulgaricusduring fermen-

tation and that enzymes were synthesized that were subsequently

useful during storage. Indeed, when catalase was added in the

products having direct inoculation of the probiotic culture in

the finished product, an increased stability during storage was

noted, while this was not the case in those where the probiotic

culture was added earlier in the manufacture (Hull et al. 1984).

It can also be argued that the gradual exposure to acid during

fermentation would also be involved in the beneficial effect of in-

oculation at the beginning of the fermentation. Thus, enzymatic

adaptation in acid environments needs to be addressed.

Protection Against Acid

Most LAB and probiotic cultures have optimum growth rates

between pH 5.5 and 6.6. Consequently, when they are exposed

for short or long periods to pH levels below 5.5 (cheese, yogurt,

fruit juices, stomach) acid diffuses into the cells and reduces

the efficiency of the enzymatic processes. Death may ultimately

result. Indeed, numerous studies report high viability loses of

probiotics exposed to the acid of the stomach (Mainville et al.

2005) as well as during storage in acid foods (Champagne et al.

2005). In the latter situation, there is often a correlation between

the poststorage pH in yogurts and the survival of probiotic bac-

teria (Kailasapathy et al. 2008).

It is well known that LAB try to maintain an intracellular

pH (pHi) constant in these acid environments, but it is not al-

ways possible. As a result, pHimay drop and generate a variety

of responses (O’Sullivan and Condon 1997). In many cases, an

acid tolerance response (ATR) occurs that significantly increases

the ability of cells to survive a short exposure (a few hours) to

high acid environments. In addition, cross-protection occurs and

ATR helps protect against short-term stresses caused by heating,

H 2 O 2 , salt, and ethanol (O’Sullivan and Condon 1997). The ef-

fect of the matrix medium on pHivaries between strains and if

affected by incubation time (Nannen and Hutkins 1991). There-

fore, ATR is not generalized amongst the LAB. Unfortunately,

most ATR studies were tested on short exposures to acidity, such

as passage through the GI tract, but little is known on the en-

zymatic factors that will improve stability to weeks of exposure

to acid. There seems to be a statistically significant correlation

between stability during storage in a fruit drink and the ability

of the strains to grow at pH 4.2, but that the relationship was not